RESUMEN
Thioredoxins (TRXs) are central to redox regulation, modulating enzyme activities to adapt metabolism to environmental changes. Previous research emphasized mitochondrial and microsomal TRX o1 and h2 influence on mitochondrial metabolism, including photorespiration and the tricarboxylic acid (TCA) cycle. Our study aimed to compare TRX-based regulation circuits towards environmental cues mainly affecting photorespiration. Metabolite snapshots, phenotypes and CO2 assimilation were compared among single and multiple TRX mutants in the wild-type and the glycine decarboxylase T-protein knockdown (gldt1) background. Our analyses provided evidence for additive negative effects of combined TRX o1 and h2 deficiency on growth and photosynthesis. Especially metabolite accumulation patterns suggest a shared regulation mechanism mainly on mitochondrial dihydrolipoamide dehydrogenase (mtLPD1)-dependent pathways. Quantification of pyridine nucleotides, in conjunction with 13C-labelling approaches, and biochemical analysis of recombinant mtLPD1 supported this. It also revealed mtLPD1 inhibition by NADH, pointing at an additional measure to fine-tune it's activity. Collectively, we propose that lack of TRX o1 and h2 perturbs the mitochondrial redox state, which impacts on other pathways through shifts in the NADH/NAD+ ratio via mtLPD1. This regulation module might represent a node for simultaneous adjustments of photorespiration, the TCA cycle and branched chain amino acid degradation under fluctuating environmental conditions.
Asunto(s)
Dihidrolipoamida Deshidrogenasa , Mitocondrias , Tiorredoxinas , Dihidrolipoamida Deshidrogenasa/metabolismo , Dihidrolipoamida Deshidrogenasa/genética , Mitocondrias/metabolismo , Tiorredoxinas/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/enzimología , Fotosíntesis , Oxidación-Reducción , NAD/metabolismo , Ambiente , Mutación , Dióxido de Carbono/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Plantas/genéticaRESUMEN
Plants contain three NADPH-thioredoxin reductases (NTR) located in the cytosol/mitochondria (NTRA/B) and the plastid (NTRC) with important metabolic functions. However, mutants deficient in all NTRs remained to be investigated. Here, we generated and characterised the triple Arabidopsis ntrabc mutant alongside with ntrc single and ntrab double mutants under different environmental conditions. Both ntrc and ntrabc mutants showed reduced growth and substantial metabolic alterations, especially in sink leaves and under high CO2 (HC), as compared to the wild type. However, ntrabc showed higher effective quantum yield of PSII under both constant and fluctuating light conditions, altered redox states of NADH/NAD+ and glutathione (GSH/GSSG) and lower potential quantum yield of PSII in sink leaves in ambient but not high CO2 concentrations, as compared to ntrc, suggesting a functional interaction between chloroplastic and extra-chloroplastic NTRs in photosynthesis regulation depending on leaf development and environmental conditions. Our results unveil a previously unknown role of the NTR system in regulating sink leaf metabolism and plant acclimation to HC, while it is not affecting full plant development, indicating that the lack of the NTR system can be compensated, at least to some extent, by other redox mechanisms.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , NADP/metabolismo , Dióxido de Carbono/metabolismo , Reductasa de Tiorredoxina-Disulfuro/genética , Reductasa de Tiorredoxina-Disulfuro/metabolismo , Arabidopsis/metabolismo , Fotosíntesis/fisiología , Cloroplastos/metabolismo , Oxidación-Reducción , Hojas de la Planta/metabolismo , Tiorredoxinas/metabolismo , AclimataciónRESUMEN
13 C-Metabolic flux analysis (13 C-MFA) has greatly contributed to our understanding of plant metabolic regulation. However, the generation of detailed in vivo flux maps remains a major challenge. Flux investigations based on nuclear magnetic resonance have resolved small networks with high accuracy. Mass spectrometry (MS) approaches have broader potential, but have hitherto been limited in their power to deduce flux information due to lack of atomic level position information. Herein we established a gas chromatography (GC) coupled to MS-based approach that provides 13 C-positional labelling information in glucose, malate and glutamate (Glu). A map of electron impact (EI)-mediated MS fragmentation was created and validated by 13 C-positionally labelled references via GC-EI-MS and GC-atmospheric pressure chemical ionization-MS technologies. The power of the approach was revealed by analysing previous 13 C-MFA data from leaves and guard cells, and 13 C-HCO3 labelling of guard cells harvested in the dark and after the dark-to-light transition. We demonstrated that the approach is applicable to established GC-EI-MS-based 13 C-MFA without the need for experimental adjustment, but will benefit in the future from paired analyses by the two GC-MS platforms. We identified specific glucose carbon atoms that are preferentially labelled by photosynthesis and gluconeogenesis, and provide an approach to investigate the phosphoenolpyruvate carboxylase (PEPc)-derived 13 C-incorporation into malate and Glu. Our results suggest that gluconeogenesis and the PEPc-mediated CO2 assimilation into malate are activated in a light-independent manner in guard cells. We further highlight that the fluxes from glycolysis and PEPc toward Glu are restricted by the mitochondrial thioredoxin system in illuminated leaves.
Asunto(s)
Carbono/análisis , Cromatografía de Gases y Espectrometría de Masas/métodos , Análisis de Flujos Metabólicos/métodos , Isótopos de Carbono/análisis , Ácido Glutámico/análisis , Glucólisis , Espectroscopía de Resonancia Magnética , Malatos/análisis , Fotosíntesis , Hojas de la Planta/metabolismoRESUMEN
Recent results suggest that metabolism-mediated stomatal closure mechanisms are important to regulate differentially the stomatal speediness between ferns and angiosperms. However, evidence directly linking mesophyll metabolism and the slower stomatal conductance (gs ) in ferns is missing. Here, we investigated the effect of exogenous application of abscisic acid (ABA), sucrose and mannitol on stomatal kinetics and carried out a metabolic fingerprinting analysis of ferns and angiosperms leaves harvested throughout a diel course. Fern stomata did not respond to ABA in the time period analysed. No differences in the relative decrease in gs was observed between ferns and the angiosperm following provision of sucrose or mannitol. However, ferns have slower gs responses to these compounds than angiosperms. Metabolomics analysis highlights that ferns have a higher accumulation of secondary rather than primary metabolites throughout the diel course, with the opposite being observed in angiosperms. Our results indicate that metabolism-mediated stomatal closure mechanisms underpin the differential stomatal speediness regulation among ferns and angiosperms, in which the slower stomatal closure in ferns is associated with the lack of ABA-responsiveness, to a reduced capacity to respond to mesophyll-derived sucrose and to a higher carbon allocation toward secondary metabolism, which likely modulates both photosynthesis-gs and growth-stress tolerance trade-offs.
Asunto(s)
Ácido Abscísico/farmacología , Helechos/fisiología , Magnoliopsida/fisiología , Manitol/farmacología , Reguladores del Crecimiento de las Plantas/farmacología , Estomas de Plantas/fisiología , Sacarosa/farmacología , Helechos/metabolismo , Cinética , Magnoliopsida/metabolismoRESUMEN
Thioredoxins (TRXs) are ubiquitous proteins engaged in the redox regulation of plant metabolism. Whilst the light-dependent TRX-mediated activation of Calvin-Benson cycle enzymes is well documented, the role of extraplastidial TRXs in the control of the mitochondrial (photo)respiratory metabolism has been revealed relatively recently. Mitochondrially located TRX o1 has been identified as a regulator of alternative oxidase, enzymes of, or associated with, the tricarboxylic acid (TCA) cycle, and the mitochondrial dihydrolipoamide dehydrogenase (mtLPD) involved in photorespiration, the TCA cycle, and the degradation of branched chain amino acids. TRXs are seemingly a major point of metabolic regulation responsible for activating photosynthesis and adjusting mitochondrial photorespiratory metabolism according to the prevailing cellular redox status. Furthermore, TRX-mediated (de)activation of TCA cycle enzymes contributes to explain the non-cyclic flux mode of operation of this cycle in illuminated leaves. Here we provide an overview on the decisive role of TRXs in the coordination of mitochondrial metabolism in the light and provide in silico evidence for other redox-regulated photorespiratory enzymes. We further discuss the consequences of mtLPD regulation beyond photorespiration and provide outstanding questions that should be addressed in future studies to improve our understanding of the role of TRXs in the regulation of central metabolism.
Asunto(s)
Arabidopsis , Arabidopsis/metabolismo , Oxidación-Reducción , Fotosíntesis , Respiración , Tiorredoxinas/metabolismoRESUMEN
Besides stomata, the photosynthetic CO2 pathway also involves the transport of CO2 from the sub-stomatal air spaces inside to the carboxylation sites in the chloroplast stroma, where Rubisco is located. This pathway is far to be a simple and direct way, formed by series of consecutive barriers that the CO2 should cross to be finally assimilated in photosynthesis, known as the mesophyll conductance (gm). Therefore, the gm reflects the pathway through different air, water and biophysical barriers within the leaf tissues and cell structures. Currently, it is known that gm can impose the same level of limitation (or even higher depending of the conditions) to photosynthesis than the wider known stomata or biochemistry. In this mini-review, we are focused on each of the gm determinants to summarize the current knowledge on the mechanisms driving gm from anatomical to metabolic and biochemical perspectives. Special attention deserve the latest studies demonstrating the importance of the molecular mechanisms driving anatomical traits as cell wall and the chloroplast surface exposed to the mesophyll airspaces (Sc/S) that significantly constrain gm. However, even considering these recent discoveries, still is poorly understood the mechanisms about signaling pathways linking the environment a/biotic stressors with gm responses. Thus, considering the main role of gm as a major driver of the CO2 availability at the carboxylation sites, future studies into these aspects will help us to understand photosynthesis responses in a global change framework.
Asunto(s)
Cloroplastos/metabolismo , Células del Mesófilo/fisiología , Fotosíntesis , Plantas/metabolismo , Ribulosa-Bifosfato Carboxilasa/fisiología , Dióxido de Carbono/fisiología , Difusión , Hojas de la Planta/metabolismo , Transducción de Señal , AguaRESUMEN
Thioredoxins (TRXs) are important proteins involved in redox regulation of metabolism. In plants, it has been shown that the mitochondrial metabolism is regulated by the mitochondrial TRX system. However, the functional significance of TRX h2, which is found at both cytosol and mitochondria, remains unclear. Arabidopsis plants lacking TRX h2 showed delayed seed germination and reduced respiration alongside impaired stomatal and mesophyll conductance, without impacting photosynthesis under ambient O2 conditions. However, an increase in the stoichiometry of photorespiratory CO2 release was found during O2 -dependent gas exchange measurements in trxh2 mutants. Metabolite profiling of trxh2 leaves revealed alterations in key metabolites of photorespiration and in several metabolites involved in respiration and amino acid metabolism. Decreased abundance of serine hydroxymethyltransferase and glycine decarboxylase (GDC) H and L subunits as well as reduced NADH/NAD+ ratios were also observed in trxh2 mutants. We further demonstrated that the redox status of GDC-L is altered in trxh2 mutants in vivo and that recombinant TRX h2 can deactivate GDC-L in vitro, indicating that this protein is redox regulated by the TRX system. Collectively, our results demonstrate that TRX h2 plays an important role in the redox regulation of mitochondrial photorespiratory metabolism.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Mitocondrias/metabolismo , Tiorredoxina h/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Dióxido de Carbono/metabolismo , Respiración de la Célula/fisiología , Clorofila A , Regulación de la Expresión Génica de las Plantas , Glicina-Deshidrogenasa (Descarboxilante)/metabolismo , Glicina Hidroximetiltransferasa , Oxidación-Reducción , Fotosíntesis/fisiología , Hojas de la Planta/metabolismo , Tiorredoxina h/genética , TranscriptomaRESUMEN
Sucrose has long been thought to play an osmolytic role in stomatal opening. However, recent evidence supports the idea that the role of sucrose in this process is primarily energetic. Here we used a combination of stomatal aperture assays and kinetic [U-13 C]-sucrose isotope labelling experiments to confirm that sucrose is degraded during light-induced stomatal opening and to define the fate of the C released from sucrose breakdown. We additionally show that addition of sucrose to the medium did not enhance light-induced stomatal opening. The isotope experiment showed a consistent 13 C enrichment in fructose and glucose, indicating that during light-induced stomatal opening sucrose is indeed degraded. We also observed a clear 13 C enrichment in glutamate and glutamine (Gln), suggesting a concerted activation of sucrose degradation, glycolysis and the tricarboxylic acid cycle. This is in contrast to the situation for Gln biosynthesis in leaves under light, which has been demonstrated to rely on previously stored C. Our results thus collectively allow us to redraw current models concerning the influence of sucrose during light-induced stomatal opening, in which, instead of being accumulated, sucrose is degraded providing C skeletons for Gln biosynthesis.
Asunto(s)
Arabidopsis/fisiología , Glutamina/biosíntesis , Glucólisis , Sacarosa/metabolismo , Arabidopsis/efectos de la radiación , Isótopos de Carbono/análisis , Ácido Glutámico/metabolismo , Ácido Glutámico/efectos de la radiación , Glutamina/efectos de la radiación , Cinética , Luz , Hojas de la Planta/fisiología , Hojas de la Planta/efectos de la radiación , Estomas de Plantas/fisiología , Estomas de Plantas/efectos de la radiación , Sacarosa/efectos de la radiaciónRESUMEN
Thioredoxins (Trxs) modulate metabolic responses during stress conditions; however, the mechanisms governing the responses of plants subjected to multiple drought events and the role of Trxs under these conditions are not well understood. Here we explored the significance of the mitochondrial Trx system in Arabidopsis following exposure to single and repeated drought events. We analyzed the previously characterized NADPH-dependent Trx reductase A and B double mutant (ntra ntrb) and two independent mitochondrial thioredoxin o1 (trxo1) mutant lines. Following similar reductions in relative water content (â¼50%), Trx mutants subjected to two drought cycles displayed a significantly higher maximum quantum efficiency (Fv/Fm) and were less sensitive to drought than their wild-type counterparts and than all genotypes subjected to a single drought event. Trx mutant plants displayed a faster recovery after two cycles of drought, as observed by the higher accumulation of secondary metabolites and higher stomatal conductance. Our results indicate that plants exposed to multiple drought cycles are able to modulate their subsequent metabolic and physiological response, suggesting the occurrence of an exquisite acclimation in stressed Arabidopsis plants. Moreover, this differential acclimation involves the participation of a set of metabolic changes as well as redox poise alteration following stress recovery.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Sequías , Mitocondrias/metabolismo , Tiorredoxinas/metabolismo , Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Metaboloma , Mutación/genética , Nucleótidos/metabolismo , Oxidación-Reducción , Estomas de Plantas/fisiología , Análisis de Componente Principal , Estrés Fisiológico , AguaRESUMEN
Stomatal responses to environmental signals differ substantially between ferns and angiosperms. However, the mechanisms that lead to such different responses remain unclear. Here we investigated the extent to which leaf metabolism contributes to coordinate the differential stomatal behaviour among ferns and angiosperms. Stomata from all species were responsive to light and CO2 transitions. However, fern stomatal responses were slower and minor in both absolute and relative terms. Angiosperms have higher stomatal density, but this is not correlated with speed of stomatal closure. The metabolic responses throughout the diel course and under different CO2 conditions differ substantially among ferns and angiosperms. Higher sucrose content and an increased sucrose-to-malate ratio during high CO2 -induced stomatal closure was observed in angiosperms compared to ferns. Furthermore, the speed of stomatal closure was positively and negatively correlated with sugars and organic acids, respectively, suggesting that the balance between sugars and organic acids aids in explaining the faster stomatal responses of angiosperms. Our results suggest that mesophyll-derived metabolic signals, especially those associated with sucrose and malate, may also be important to modulate the differential stomatal behaviour between ferns and angiosperms, providing important new information that helps in understanding the metabolism-mediated mechanisms regulating stomatal movements across land plant evolution.
Asunto(s)
Dióxido de Carbono/metabolismo , Helechos/fisiología , Luz , Magnoliopsida/fisiología , Malatos/metabolismo , Estomas de Plantas/metabolismo , Estomas de Plantas/efectos de la radiación , Sacarosa/metabolismo , Análisis Discriminante , Helechos/efectos de la radiación , Análisis de los Mínimos Cuadrados , Magnoliopsida/efectos de la radiación , Metaboloma/efectos de la radiación , Fotosíntesis/efectos de la radiación , Análisis de Componente PrincipalRESUMEN
Auxin modulates a range of plant developmental processes including embryogenesis, organogenesis, and shoot and root development. Recent studies have shown that plant hormones also strongly influence metabolic networks, which results in altered growth phenotypes. Modulating auxin signalling pathways may therefore provide an opportunity to alter crop performance. Here, we performed a detailed physiological and metabolic characterization of tomato (Solanum lycopersicum) mutants with either increased (entire) or reduced (diageotropica-dgt) auxin signalling to investigate the consequences of altered auxin signalling on photosynthesis, water use, and primary metabolism. We show that reduced auxin sensitivity in dgt led to anatomical and physiological modifications, including altered stomatal distribution along the leaf blade and reduced stomatal conductance, resulting in clear reductions in both photosynthesis and water loss in detached leaves. By contrast, plants with higher auxin sensitivity (entire) increased the photosynthetic capacity, as deduced by higher Vcmax and Jmax coupled with reduced stomatal limitation. Remarkably, our results demonstrate that auxin-sensitive mutants (dgt) are characterized by impairments in the usage of starch that led to lower growth, most likely associated with decreased respiration. Collectively, our findings suggest that mutations in different components of the auxin signalling pathway specifically modulate photosynthetic and respiratory processes.
Asunto(s)
Ácidos Indolacéticos/metabolismo , Mitocondrias/metabolismo , Fotosíntesis/fisiología , Reguladores del Crecimiento de las Plantas/metabolismo , Transducción de Señal , Solanum lycopersicum/crecimiento & desarrollo , Clorofila/metabolismo , Solanum lycopersicum/metabolismo , Solanum lycopersicum/fisiología , Hojas de la Planta/anatomía & histología , Estomas de Plantas/fisiología , Transducción de Señal/fisiología , Agua/metabolismoRESUMEN
Retrograde signalling pathways that are triggered by changes in cellular redox homeostasis remain poorly understood. Transformed rice plants that are deficient in peroxisomal ascorbate peroxidase APX4 (OsAPX4-RNAi) are known to exhibit more effective protection of photosynthesis against oxidative stress than controls when catalase (CAT) is inhibited, but the mechanisms involved have not been characterized. An in-depth physiological and proteomics analysis was therefore performed on OsAPX4-RNAi CAT-inhibited rice plants. Loss of APX4 function led to an increased abundance of several proteins that are involved in essential metabolic pathways, possibly as a result of increased tissue H2O2 levels. Higher photosynthetic activities observed in the OsAPX4-RNAi plants under CAT inhibition were accompanied by higher levels of Rubisco, higher maximum rates of Rubisco carboxylation, and increased photochemical efficiencies, together with large increases in photosynthesis-related proteins. Large increases were also observed in the levels of proteins involved in the ascorbate/glutathione cycle and in other antioxidant-related pathways, and these changes may be important in the protection of photosynthesis in the OsAPX4-RNAi plants. Large increases in the abundance of proteins localized in the nuclei and mitochondria were also observed, together with increased levels of proteins involved in important cellular pathways, particularly protein translation. Taken together, the results show that OsAPX4-RNAi plants exhibit significant metabolic reprogramming, which incorporates a more effective antioxidant response to protect photosynthesis under conditions of impaired CAT activity.
Asunto(s)
Ascorbato Peroxidasas/metabolismo , Catalasa/metabolismo , Oryza/metabolismo , Estrés Oxidativo , Fotosíntesis , Interferencia de ARNRESUMEN
Malate is a central metabolite involved in a multiplicity of plant metabolic pathways, being associated with mitochondrial metabolism and playing significant roles in stomatal movements. Vacuolar malate transport has been characterized at the molecular level and is performed by at least one carrier protein and two channels in Arabidopsis (Arabidopsis thaliana) vacuoles. The absence of the Arabidopsis tonoplast Dicarboxylate Transporter (tDT) in the tdt knockout mutant was associated previously with an impaired accumulation of malate and fumarate in leaves. Here, we investigated the consequences of this lower accumulation on stomatal behavior and photosynthetic capacity as well as its putative metabolic impacts. Neither the stomatal conductance nor the kinetic responses to dark, light, or high CO2 were highly affected in tdt plants. In addition, we did not observe any impact on stomatal aperture following incubation with abscisic acid, malate, or citrate. Furthermore, an effect on photosynthetic capacity was not observed in the mutant lines. However, leaf mitochondrial metabolism was affected in the tdt plants. Levels of the intermediates of the tricarboxylic acid cycle were altered, and increases in both light and dark respiration were observed. We conclude that manipulation of the tonoplastic organic acid transporter impacted mitochondrial metabolism, while the overall stomatal and photosynthetic capacity were unaffected.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/metabolismo , Fumaratos/metabolismo , Malatos/metabolismo , Mutación/genética , Transportadores de Anión Orgánico/genética , Estomas de Plantas/fisiología , Vacuolas/metabolismo , Aminoácidos/metabolismo , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/metabolismo , Respiración de la Célula , Clorofila/metabolismo , Clorofila A , Ciclo del Ácido Cítrico , Fluorescencia , Técnicas de Inactivación de Genes , Metaboloma , Transportadores de Anión Orgánico/metabolismo , Fotoperiodo , Fotosíntesis , Estomas de Plantas/citología , Almidón/metabolismoRESUMEN
Plant mitochondria have a fully operational tricarboxylic acid (TCA) cycle that plays a central role in generating ATP and providing carbon skeletons for a range of biosynthetic processes in both heterotrophic and photosynthetic tissues. The cycle enzyme-encoding genes have been well characterized in terms of transcriptional and effector-mediated regulation and have also been subjected to reverse genetic analysis. However, despite this wealth of attention, a central question remains unanswered: "What regulates flux through this pathway in vivo?" Previous proteomic experiments with Arabidopsis discussed below have revealed that a number of mitochondrial enzymes, including members of the TCA cycle and affiliated pathways, harbor thioredoxin (TRX)-binding sites and are potentially redox-regulated. We have followed up on this possibility and found TRX to be a redox-sensitive mediator of TCA cycle flux. In this investigation, we first characterized, at the enzyme and metabolite levels, mutants of the mitochondrial TRX pathway in Arabidopsis: the NADP-TRX reductase a and b double mutant (ntra ntrb) and the mitochondrially located thioredoxin o1 (trxo1) mutant. These studies were followed by a comparative evaluation of the redistribution of isotopes when (13)C-glucose, (13)C-malate, or (13)C-pyruvate was provided as a substrate to leaves of mutant or WT plants. In a complementary approach, we evaluated the in vitro activities of a range of TCA cycle and associated enzymes under varying redox states. The combined dataset suggests that TRX may deactivate both mitochondrial succinate dehydrogenase and fumarase and activate the cytosolic ATP-citrate lyase in vivo, acting as a direct regulator of carbon flow through the TCA cycle and providing a mechanism for the coordination of cellular function.
Asunto(s)
Ciclo del Ácido Cítrico , Mitocondrias/metabolismo , Tiorredoxinas/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Isótopos de Carbono , Citratos/metabolismo , Genes de Plantas , Prueba de Complementación Genética , Metabolómica , Modelos Biológicos , Mutación/genética , Hojas de la Planta/enzimología , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , Plastidios/metabolismo , Reproducibilidad de los Resultados , Semillas/crecimiento & desarrollo , Semillas/metabolismoRESUMEN
Contents 1018 I. 1018 II. 1019 III. 1022 IV. 1025 V. 1026 VI. 1029 1030 References 1030 SUMMARY: Stomata are leaf epidermal structures consisting of two guard cells surrounding a pore. Changes in the aperture of this pore regulate plant water-use efficiency, defined as gain of C by photosynthesis per leaf water transpired. Stomatal aperture is actively regulated by reversible changes in guard cell osmolyte content. Despite the fact that guard cells can photosynthesize on their own, the accumulation of mesophyll-derived metabolites can seemingly act as signals which contribute to the regulation of stomatal movement. It has been shown that malate can act as a signalling molecule and a counter-ion of potassium, a well-established osmolyte that accumulates in the vacuole of guard cells during stomatal opening. By contrast, their efflux from guard cells is an important mechanism during stomatal closure. It has been hypothesized that the breakdown of starch, sucrose and lipids is an important mechanism during stomatal opening, which may be related to ATP production through glycolysis and mitochondrial metabolism, and/or accumulation of osmolytes such as sugars and malate. However, experimental evidence supporting this theory is lacking. Here we highlight the particularities of guard cell metabolism and discuss this in the context of the guard cells themselves and their interaction with the mesophyll cells.
Asunto(s)
Fotosíntesis , Estomas de Plantas/fisiología , Metabolismo de los Hidratos de Carbono , Células del Mesófilo/metabolismo , Almidón/metabolismoRESUMEN
Stomata control the exchange of CO2 and water vapor in land plants. Thus, whereas a constant supply of CO2 is required to maintain adequate rates of photosynthesis, the accompanying water losses must be tightly regulated to prevent dehydration and undesired metabolic changes. Accordingly, the uptake or release of ions and metabolites from guard cells is necessary to achieve normal stomatal function. The AtQUAC1, an R-type anion channel responsible for the release of malate from guard cells, is essential for efficient stomatal closure. Here, we demonstrate that mutant plants lacking AtQUAC1 accumulated higher levels of malate and fumarate. These mutant plants not only display slower stomatal closure in response to increased CO2 concentration and dark but are also characterized by improved mesophyll conductance. These responses were accompanied by increases in both photosynthesis and respiration rates, without affecting the activity of photosynthetic and respiratory enzymes and the expression of other transporter genes in guard cells, which ultimately led to improved growth. Collectively, our results highlight that the transport of organic acids plays a key role in plant cell metabolism and demonstrate that AtQUAC1 reduce diffusive limitations to photosynthesis, which, at least partially, explain the observed increments in growth under well-watered conditions.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiología , Células del Mesófilo/fisiología , Transportadores de Anión Orgánico/metabolismo , Fotosíntesis/fisiología , Estomas de Plantas/fisiología , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Carbono/metabolismo , Dióxido de Carbono/metabolismo , Sequías , Enzimas/genética , Enzimas/metabolismo , Fumaratos/metabolismo , Regulación de la Expresión Génica de las Plantas , Técnicas de Inactivación de Genes , Malatos/metabolismo , Mutación , Transportadores de Anión Orgánico/genética , Hojas de la Planta/genética , Hojas de la Planta/crecimiento & desarrollo , Estomas de Plantas/genéticaRESUMEN
The control of stomatal aperture involves reversible changes in the concentration of osmolytes in guard cells. Sucrose has long been proposed to have an osmolytic role in guard cells. However, direct evidence for such a role is lacking. Furthermore, recent evidence suggests that sucrose may perform additional roles in guard cells. Here, we provide an update covering the multiple roles of sucrose in guard cell regulation, highlighting the knowledge accumulated regarding spatiotemporal differences in the synthesis, accumulation, and degradation of sucrose as well as reviewing the role of sucrose as a metabolic connector between mesophyll and guard cells. Analysis of transcriptomic data from previous studies reveals that several genes encoding sucrose and hexose transporters and genes involved in gluconeogenesis, sucrose and trehalose metabolism are highly expressed in guard cells compared with mesophyll cells. Interestingly, this analysis also showed that guard cells have considerably higher expression of C4 -marker genes than mesophyll cells. We discuss the possible roles of these genes in guard cell function and the role of sucrose in stomatal opening and closure. Finally, we provide a perspective for future experiments which are required to fill gaps in our understanding of both guard cell metabolism and stomatal regulation.
Asunto(s)
Estomas de Plantas/citología , Estomas de Plantas/metabolismo , Sacarosa/metabolismo , Células del Mesófilo/metabolismo , Ósmosis , Fotosíntesis , Almidón/metabolismoRESUMEN
Isoform 3 of sucrose synthase (SUS3) is highly expressed in guard cells; however, the precise function of SUS3 in this cell type remains to be elucidated. Here, we characterized transgenic Nicotiana tabacum plants overexpressing SUS3 under the control of the stomatal-specific KST1 promoter, and investigated the changes in guard cell metabolism during the dark to light transition. Guard cell-specific SUS3 overexpression led to increased SUS activity, stomatal aperture, stomatal conductance, transpiration rate, net photosynthetic rate and growth. Although only minor changes were observed in the metabolite profile in whole leaves, an increased fructose level and decreased organic acid levels and sucrose to fructose ratio were observed in guard cells of transgenic lines. Furthermore, guard cell sucrose content was lower during light-induced stomatal opening. In a complementary approach, we incubated guard cell-enriched epidermal fragments in (13) C-NaHCO3 and followed the redistribution of label during dark to light transitions; this revealed increased labeling in metabolites of, or associated with, the tricarboxylic acid cycle. The results suggest that sucrose breakdown is a mechanism to provide substrate for the provision of organic acids for respiration, and imply that manipulation of guard cell metabolism may represent an effective strategy for plant growth improvement.
Asunto(s)
Glucosiltransferasas/metabolismo , Proteínas de Plantas/metabolismo , Estomas de Plantas/citología , Solanum tuberosum/enzimología , Sacarosa/metabolismo , Regulación hacia Arriba , Adaptación Fisiológica , Isótopos de Carbono , Ácidos Carboxílicos/metabolismo , Sequías , Gases/metabolismo , Glucosiltransferasas/genética , Cinética , Luz , Metaboloma , Metabolómica , Especificidad de Órganos , Fenotipo , Desarrollo de la Planta , Hojas de la Planta/metabolismo , Proteínas de Plantas/genética , Transpiración de Plantas/fisiología , Plantas Modificadas Genéticamente , Regiones Promotoras Genéticas/genética , Nicotiana/genéticaRESUMEN
During dark-induced senescence isovaleryl-CoA dehydrogenase (IVDH) and D-2-hydroxyglutarate dehydrogenase (D-2HGDH) act as alternate electron donors to the ubiquinol pool via the electron-transfer flavoprotein/electron-transfer flavoprotein:ubiquinone oxidoreductase (ETF/ETFQO) pathway. However, the role of this pathway in response to other stresses still remains unclear. Here, we demonstrated that this alternative pathway is associated with tolerance to drought in Arabidopsis. In comparison with wild type (WT) and lines overexpressing D-2GHDH, loss-of-function etfqo-1, d2hgdh-2 and ivdh-1 mutants displayed compromised respiration rates and were more sensitive to drought. Our results demonstrated that an operational ETF/ETFQO pathway is associated with plants' ability to withstand drought and to recover growth once water becomes replete. Drought-induced metabolic reprogramming resulted in an increase in tricarboxylic acid (TCA) cycle intermediates and total amino acid levels, as well as decreases in protein, starch and nitrate contents. The enhanced levels of the branched-chain amino acids in loss-of-function mutants appear to be related to their increased utilization as substrates for the TCA cycle under water stress. Our results thus show that mitochondrial metabolism is highly active during drought stress responses and provide support for a role of alternative respiratory pathways within this response.